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iPSCs and Gene Correction Technology Team up to Tackle Beta Thalassemia

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Review of “Enhancement of b-Globin Gene Expression in Thalassemic IVS2-654 Induced Pluripotent Stem Cell-Derived Erythroid Cells by Modified U7 snRNA” from Stem Cells Translational Medicine by Stuart P. Atkinson

Therapeutic strategies combining patient-specific induced pluripotent stem cell (iPSC) technology with genetic correction strategies have the potential to tackle a wide variety of diseases, including the beta thalassemia group of inherited blood disorders. 

In one form of beta thalassemia prevalent in Thailand and China, a specific mutation (IVS2-654) affecting the second intron of the beta hemoglobin gene (HBB) affects pre-mRNA splicing and prevents the production of functional -globin protein [1, 2]. One possible means to tackle this problem is the application of synthetic splice-switching oligonucleotides (SSOs) in patient-derived iPSCs to restore correctly spliced beta globin mRNA. Subsequent differentiation and transplantation of hematopoietic stem cells (HSCs) from corrected iPSCs may then serve as a patient-specific treatment for this blood disorder.

In a new Stem Cells Translational Medicine study, researchers from the laboratory of Suradej Hongeng (Mahidol University, Nakhon Pathom, Thailand) now report on two key advances towards a viable beta thalassemia treatment [3]. Phanthong et al describe, for the first time, the generation of iPSCs from a compound heterozygous IVS2-654 beta thalassemia patient and the correction of the splicing defect using a U7 small nuclear (sn)RNA modified to carry a splice-switching sequence [4, 5]. 

Can iPSCs and gene correction technology team up to tackle beta thalassemia in human patients?

The study generated iPSCs from bone marrow-derived mesenchymal stem cells (MSCs) from a healthy donor and a heterozygous mutation carrying-patient using a polycistronic lentivirus harboring the OCT4, SOX2, KLF4, SOX2, and c-MYC reprogramming factors. The authors then stably transfected the mutant iPSCs with a modified U7 snRNA targeted to the mutated pre-mRNA and differentiated the resultant clones into erythroblasts (nucleated red blood cells). The healthy and corrected iPSCs underwent differentiation in a similar manner and, encouragingly, the corrected iPSCs expressed correctly spliced beta globin mRNA at a level equivalent to 80% of the healthy control (See Figure).

These exciting results suggest that the transplantation of HSCs derived from U7 snRNA corrected patient-specific iPSC into beta thalassemia patients may represent an exciting means to provide levels of correctly spliced beta globin mRNA sufficient to inhibit disease symptoms. While obstacles to this goal remain, the application of the modified U7 snRNA proved to a notable success when compared to the application of other gene editing technologies [6] and supports further development of this strategy. Furthermore, the heterozygous IVS2-654 iPSCs may also prove to be a useful tool for disease modeling and drug screening purposes.

The signs look good, so stay tuned to the Stem Cells Portal and Stem Cells Translational Medicine to hear more on how iPSCs and gene correction technology can team up to tackle beta thalassemia!

References

  1. Zhang JZ, Cai SP, He X, et al. Molecular basis of beta thalassemia in south China. Strategy for DNA analysis. Hum Genet 1988;78:37-40.
  2. Huang SZ, Zeng FY, Ren ZR, et al. RNA transcripts of the beta-thalassaemia allele IVS-2-654 C-->T: a small amount of normally processed beta-globin mRNA is still produced from the mutant gene. Br J Haematol 1994;88:541-546.
  3. Phanthong P, Borwornpinyo S, Kitiyanant N, et al. Enhancement of β-Globin Gene Expression in Thalassemic IVS2-654 Induced Pluripotent Stem Cell-Derived Erythroid Cells by Modified U7 snRNA. STEM CELLS Translational Medicine 2017, In Press.
  4. Gorman L, Mercatante DR, and Kole R. Restoration of correct splicing of thalassemic beta-globin pre-mRNA by modified U1 snRNAs. J Biol Chem 2000;275:35914-35919.
  5. Vacek MM, Ma H, Gemignani F, et al. High-level expression of hemoglobin A in human thalassemic erythroid progenitor cells following lentiviral vector delivery of an antisense snRNA. Blood 2003;101:104-111.
  6. Xu P, Tong Y, Liu XZ, et al. Both TALENs and CRISPR/Cas9 directly target the HBB IVS2-654 (C > T) mutation in beta-thalassemia-derived iPSCs. Sci Rep 2015;5:12065.